updated 6:41 p.m. ET April 30, 2009

LOUISVILLE, Ky. - The biggest change at this year’s Kentucky Derby won’t be noticed by any fan or disrupt the routine of any horse. In fact, its only evidence will be sealed and stored in a padlocked refrigerator minutes after the race.

For the first time, the signature American thoroughbred race is screening for anabolic steroids — a quiet step that industry officials are hoping will make a loud splash in public relations.

“Our existence depends on public confidence,” longtime breeder Arthur Hancock said. “If we lose that, we lose our livelihood. Its extremely important we get this mess cleaned up.”

Last year’s Derby winner, Big Brown, was on steroids at the time of his victory — a fact known only because trainer Rick Dutrow acknowledged it. Dutrow explained he injected the horse with regular doses of the then-legal steroid stanozolol, sold under the brand name Winstrol, although he insisted the intent was not to build muscle but to increase his appetite and brighten his coat.

If the winner of this year’s Derby tests positive for more than a trace amount of stanozolol, the horse will be disqualified and the trainer will be subjected to a lengthy suspension.

That drug is now allowed in the sport only for therapeutic uses, and no shot can be given in Kentucky within 60 days of a race — more than enough time for any performance-enhancing effect to wear off. Only three other anabolic steroids are allowed even in minuscule doses anymore. Dozens of others have been banned completely.

Video   CNBC's Run for the Roses A look at the business of horse racing in the state of Kentucky. "Run for the Roses" premieres Thursday @ 9PM ET. NBC Sports

At the time of last year’s Derby, only 12 of 38 racing states had banned steroids. Now 35 have, representing more than 99 percent of the races subjected to pari-mutuel betting, according to the Racing Medication and Testing Consortium, an industry group that pushed a national testing standard.

“We went from completely unregulated to uniformly regulated in a year and a half,” said Scot Waterman, RMTC’s executive director. “There are plenty of people in this industry that never thought they’d see that day.”

Hall of Fame trainer Jack Van Berg compared horse training to “chemical warfare” last year while testifying to Congress on safety problems with the sport. While few in the industry are using that kind of terminology, there has been some evidence steroids were the rule, not the exception, for the 1,200-pound athletes.

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Pennsylvania banned anabolic steroids last April but had previously conducted tests to see how many of its racehorses would have tested positive. Waterman said the results were staggering — showing nearly two-thirds of the horses tested had a noticeable concentration of the drugs in their bloodstream.

Those numbers don’t surprise Larry Jones, who trained last year’s Derby runner-up Eight Belles. The filly pulled up lame jogging past the finish and had to be euthanized with two fractured ankles. With speculation swirling the muscular horse was using steroids, Jones called for not just the regular necropsy but a more sophisticated one that included a drug test to prove she was clean.

“Every wrongdoing you could do to a horse, we had been accused of it,” Jones said. “We were probably the only one of the 20 (Derby starters) not on steroids.”

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